Original article Minimized cell usage for stem cell-derived and primary cells on an automated patch clamp system Nadine Becker a, ⁎, Sonja Stoelzle a , Sven Göpel b , David Guinot a , Patrick Mumm a , Claudia Haarmann a , Daniela Malan c , Heribert Bohlen d , Eugen Kossolov d , Ralf Kettenhofen d , Michael George a , Niels Fertig a , Andrea Brüggemann a a Nanion Technologies GmbH, Gabrielenstraße 9, 80636 Munich, Germany b AstraZeneca R&D, Pepparedsleden 1, 43183 Mölndal, Sweden c Institute of Physiology I, University of Bonn Life & Brain, 53105 Bonn, Germany d Axiogenesis AG, Nattermannallee 1/S20, 50829 Cologne, Germany abstract article info Article history: 8 February 2013 27 March 2013 Available online xxxx Keywords: Automated patch clamp stem cell-derived cardiomyocytes stem cell-derived neurons pancreatic islet cells primary cells Patchliner Introduction: Chip-based automated patch clamp systems are widely used in drug development and safety pharmacology, allowing for high quality, high throughput screening at standardized experimental condi- tions. The merits of automation generally come at the cost of large amounts of cells needed, since cells are not targeted individually, but randomly positioned onto the chip aperture from cells in suspension. While cell usage is of little concern when using standard cell lines such as CHO or HEK cells, it becomes a crucial constraint with cells of limited availability, such as primary or otherwise rare and expensive cells, like in- duced pluripotent stem (IPS) cell-derived cardiomyocytes or neurons. Methods: We established application protocols for CHO cells, IPS cell-derived neurons (iCell® Neurons, Cellular Dynamics International), cardiomyocytes (Cor.4U®, Axiogenesis) and pancreatic islet cells, minimizing cell usage for automated patch clamp recordings on Nanion's Patchliner. Use of 5 μl cell suspension per well for den- sities between 55,000 cells/ml and 400,000 cells/ml depending on cell type resulted in good cell capture. Results: We present a new cell application procedure optimized for the Patchliner achieving > 80% success rates for using as little as 300 to 2000 cells per well depending on cell type. We demonstrate that this protocol works for standard cell lines, as well as for stem cell-derived neurons and cardiomyocytes, and for primary pancreatic islet cells. We present recordings for these cell types, demonstrating that high data quality is not compromised by altered cell application. Discussion: Our new cell application procedure achieves high success rates with unprecedentedly low cell num- bers. Compared to other standard automated patch clamp systems we reduced the average amount of cells need- ed by more than 150 times. Reduced cell usage crucially improves cost efficiency for expensive cells and opens up automated patch clamp for primary cells of limited availability. © 2013 Elsevier Inc. All rights reserved. 1. Introduction In recent years, automated patch clamp systems have become an indispensable asset in drug development and safety pharmacology (Möller & Witchel, 2011; Polonchuk, 2012) and led to a significant in- crease in efficiency in basic and applied research (Milligan et al., 2009). Automation of the experimental procedure and parallel pro- cessing of currently up to 384 single cell measurements allow for high throughput and standardized experimental conditions com- pared to manual patch clamp. This progress has become possible through the advent of planar patch clamp, in which the glass pipette used to approach cells in conventional patch clamp is replaced by a planar glass chip with a micrometer-sized hole. Instead of selecting and targeting specific cells with a movable pipette, planar patch clamp is based on flushing a suspension of cells over a static chip, with cells being caught by negative pressure applied through the ap- ertures in the recording chip (Milligan et al., 2009). Although this ap- proach overcomes former limitations of a tedious manual procedure requiring extensive practical training, it bears the disadvantage that a substantial amount of cells is irretrievably lost with each attempt to patch a single cell. The success rate of catching a cell for recording critically depends on the volume and density of the cell suspension flushed over each chip aperture. While the amount of cells needed is usually not a constraint for studying ion channels in standard cell lines like CHO or HEK cell lines, it is critically restricting the use of pri- mary cells derived from limited amounts of tissue or cells designed and produced at high cost, such as induced pluripotent stem (IPS) Journal of Pharmacological and Toxicological Methods xxx (2013) xxx–xxx ⁎ Corresponding author. Tel.: +49 2189 97969. E-mail address: nadine@nanion.de (N. Becker). JPM-06117; No of Pages 6 1056-8719/$ – see front matter © 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.vascn.2013.03.009 Contents lists available at SciVerse ScienceDirect Journal of Pharmacological and Toxicological Methods journal homepage: www.elsevier.com/locate/jpharmtox Please cite this article as: Becker, N., et al., Minimized cell usage for stem cell-derived and primary cells on an automated patch clamp system, Journal of Pharmacological and Toxicological Methods (2013), http://dx.doi.org/10.1016/j.vascn.2013.03.009